High-throughput quantification of antibiotic resistance genes from an urban wastewater treatment plant

Karkman, Antti; Johnson, Timothy A.; Lyra, Christina; Stedtfeld, Robert D.; Tamminen, Manu; Tiedje, James M.; Virta, Marko

doi:10.1093/femsec/fiw014

Cited by 178 publications

(85 citation statements)

References 27 publications

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“…PCR cycling conditions and initial data processing were as previously described (Wang et al, 2014). However, the detection limit was at a threshold cycle (C T ) of 27 (Zhu et al, 2013; Karkman et al, 2016; Muziasari et al, 2016). The melting curve of each PCR product was also analyzed to monitor the specificity of the primer sets.…”

Section: Methodsmentioning

confidence: 99%

“…Highly parallel qPCR arrays provide a culture-independent method that permits the combination of hundreds of assays to detect and quantify selected known genes in a single experiment. This method has been used to analyze the resistome composition in environmental samples (Looft et al, 2012; Zhu et al, 2013; Wang et al, 2014; Su et al, 2015; Karkman et al, 2016) and fish farming environments (Muziasari et al, 2016). …”

Section: Introductionmentioning

confidence: 99%

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The Resistome of Farmed Fish Feces Contributes to the Enrichment of Antibiotic Resistance Genes in Sediments below Baltic Sea Fish Farms

et al. 2017

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Our previous studies showed that particular antibiotic resistance genes (ARGs) were enriched locally in sediments below fish farms in the Northern Baltic Sea, Finland, even when the selection pressure from antibiotics was negligible. We assumed that a constant influx of farmed fish feces could be the plausible source of the ARGs enriched in the farm sediments. In the present study, we analyzed the composition of the antibiotic resistome from the intestinal contents of 20 fish from the Baltic Sea farms. We used a high-throughput method, WaferGen qPCR array with 364 primer sets to detect and quantify ARGs, mobile genetic elements (MGE), and the 16S rRNA gene. Despite a considerably wide selection of qPCR primer sets, only 28 genes were detected in the intestinal contents. The detected genes were ARGs encoding resistance to sulfonamide (sul1), trimethoprim (dfrA1), tetracycline [tet(32), tetM, tetO, tetW], aminoglycoside (aadA1, aadA2), chloramphenicol (catA1), and efflux-pumps resistance genes (emrB, matA, mefA, msrA). The detected genes also included class 1 integron-associated genes (intI1, qacEΔ1) and transposases (tnpA). Importantly, most of the detected genes were the same genes enriched in the farm sediments. This preliminary study suggests that feces from farmed fish contribute to the ARG enrichment in farm sediments despite the lack of contemporaneous antibiotic treatments at the farms. We observed that the intestinal contents of individual farmed fish had their own resistome compositions. Our result also showed that the total relative abundances of transposases and tet genes were significantly correlated (p = 0.001, R2 = 0.71). In addition, we analyzed the mucosal skin and gill filament resistomes of the farmed fish but only one multidrug-efflux resistance gene (emrB) was detected. To our knowledge, this is the first study reporting the resistome of farmed fish using a culture-independent method. Determining the possible sources of ARGs, especially mobilized ARGs, is essential for controlling the occurrence and spread of ARGs at fish farming facilities and for lowering the risk of ARG spread from the farms to surrounding environments.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

The Resistome of Farmed Fish Feces Contributes to the Enrichment of Antibiotic Resistance Genes in Sediments below Baltic Sea Fish Farms

et al. 2017

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“…In the absence of access to any data on the type or rate of antibiotic prescription in the community, we decided to look for a subset of genes that have previously been shown to be commonly found in various aquatic environments (35,36). A two-pronged approach to detect various antibiotic resistance genes, a qPCR…”

Section: Discussionmentioning

confidence: 99%

Detection of Antibiotic Resistance Genes in Source and Drinking Water Samples from a First Nations Community in Canada

Fernando

Tun

Poole

et al. 2016

Appl Environ Microbiol

104

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Access to safe drinking water is now recognized as a human right by the United Nations. In developed countries like Canada, access to clean water is generally not a matter of concern. However, one in every five First Nations reserves is under a drinking water advisory, often due to unacceptable microbiological quality. In this study, we analyzed source and potable water from a First Nations community for the presence of coliform bacteria as well as various antibiotic resistance genes. Samples, including those from drinking water sources, were found to be positive for various antibiotic resistance genes, namely, ampC, tet(A), mecA, ␤-lactamase genes (SHV-type, TEM-type, CTX-M-type, OXA-1, and CMY-2-type), and carbapenemase genes (KPC, IMP, VIM, NDM, GES, and OXA-48 genes). Not surprisingly, substantial numbers of total coliforms, including Escherichia coli, were recovered from these samples, and this result was also confirmed using Illumina sequencing of the 16S rRNA gene. These findings deserve further attention, as the presence of coliforms and antibiotic resistance genes potentially puts the health of the community members at risk. IMPORTANCEIn this study, we highlight the poor microbiological quality of drinking water in a First Nations community in Canada. We examined the coliform load as well as the presence of antibiotic resistance genes in these samples. This study examined the presence of antibiotic-resistant genes in drinking water samples from a First Nations Community in Canada. We believe that our findings are of considerable significance, since the issue of poor water quality in First Nations communities in Canada is often ignored, and our findings will help shed some light on this important issue.A ntibiotic resistance in bacteria has been recognized as one of the greatest threats to human health by the World Health Organization (1). Overuse and misuse of antibiotics contribute to the buildup of selective pressure aiding the proliferation of antibiotic-resistant bacteria (2, 3). While hospital environments are notorious for selecting for antibiotic-resistant bacteria, it is now becoming increasingly evident that overuse and misuse of antibiotics are also creating a selective pressure outside hospital settings. Studies over the last few years have shown the presence of antibiotics and of antibiotic-resistant bacteria in the broader environment, including water supplies and soil samples (4). This is indeed alarming as the high number of antibiotic-resistant bacteria in communities makes the treatment of community-acquired infections increasingly challenging (5, 6).Not surprisingly, water samples from communities that lack access to clean water contain high numbers of bacteria (7-9). While a high bacterial count in the water supply itself poses an increased health risk (10), the presence of antibiotic-resistant bacteria makes this risk even more serious. Lack of access to clean and safe water is a problem that is generally associated with developing countries; however, this is a reality as wel...

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“…A wide variety of existing molecular techniques have been demonstrated to recover and analyse environmental metagenomic DNA [15,16,44]. These techniques include next-generation sequencing as well as chip-based methodologies, though perhaps the simplest and most cost-effective methods use quantitative PCR (QPCR).…”

Section: Methodsmentioning

confidence: 99%

The antimicrobial resistance crisis: management through gene monitoring

2016

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Antimicrobial resistance (AMR) is an acknowledged crisis for humanity. Its genetic origins and dire potential outcomes are increasingly well understood. However, diagnostic techniques for monitoring the crisis are currently largely limited to enumerating the increasing incidence of resistant pathogens. Being the end-stage of the evolutionary process that produces antimicrobial resistant pathogens, these measurements, while diagnostic, are not prognostic, and so are not optimal in managing this crisis. A better test is required. Here, using insights from an understanding of evolutionary processes ruling the changing abundance of genes under selective pressure, we suggest a predictive framework for the AMR crisis. We then discuss the likely progression of resistance for both existing and prospective antimicrobial therapies. Finally, we suggest that by the environmental monitoring of resistance gene frequency, resistance may be detected and tracked presumptively, and how this tool may be used to guide decision-making in the local and global use of antimicrobials.

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High-throughput quantification of antibiotic resistance genes from an urban wastewater treatment plant

Cited by 178 publications

References 27 publications

The Resistome of Farmed Fish Feces Contributes to the Enrichment of Antibiotic Resistance Genes in Sediments below Baltic Sea Fish Farms

The Resistome of Farmed Fish Feces Contributes to the Enrichment of Antibiotic Resistance Genes in Sediments below Baltic Sea Fish Farms

Detection of Antibiotic Resistance Genes in Source and Drinking Water Samples from a First Nations Community in Canada

The antimicrobial resistance crisis: management through gene monitoring

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